G protein-gated inwardly rectifying K+ (GIRK) channels were the first example of an effector that is directly regulated by the beta-gamma subunits of G proteins. However, relatively little is known about the elements on the Gbeta-gamma that are important for GIRK channel regulation. We have previously used a chimeric strategy to identify important elements on the channel that are important for regulation by Gbeta-gamma (He et. al. 1999). Here, we present preliminary data indicating that in Xenopus oocytes, Gbeta1-beta4 can all activate the GIRK channels with similar efficacy. However, the Gbeta5 is incapable of activating these channels under identical conditions. Western blot analysis in oocytes expressing Gbeta5 showed that the protein is expressed. Therefore lack of expression can not be responsible for lack of channel activation. Sequence alignment showed that Gbeta1-beta4 display approximately 90% identity, whereas Gbeta5 shows only 51 % identity with other Gbeta subunits. Based on these findings, we propose to use a chimeric strategy between Gbeta1 and Gbeta5 to identify the regions responsible for the difference in channel activation by the two subunits. Once these regions are identified, we will use mutagenesis to identify specific residues involved in the interaction of Gbeta1 and the channel. These studies will help us better understand the interaction of G proteins with GIRK channels. The proposed studies will also help us achieve a better understanding of G protein mediate signaling, which is of major physiological importance.